Date of Graduation
Retroviruses are a family of RNA viruses that replicate through DNA intermediates by reverse transcribing their genome. One of the essential steps of reverse transcription is minus-strand DNA transfer, during which nascent DNA switches template usage from the 5â€² end to near the 3â€² of the viral RNA genome. We have determined that minus-strand DNA transfer is homology driven; a minimum homology length is required to align the newly synthesized DNA with the 3â€² RNA template to facilitate an efficient and accurate transfer. In addition to the obligatory template strand transfer events necessary to complete reverse transcription, template switching also occurs to generate recombinants by using the two co-packaged RNA as template during DNA synthesis. In order for observable recombination events to occur, a virion must have two genetically different RNA (heterozygote). Heterozygotic virions are produced by a cell infected with two genetically distinct viruses. Although an important factor for viral variation, very little is known about the nature of double infection. We investigated the dynamics of double infection (DI) and found that retroviral double infection occurs more frequently than it would at random. We observed that human immunodeficiency virus type 1 HIV-1) DI is not random in cultured T cells and in primary T cells. Moreover, DI is not random by direct and by cell-mediated transmission of HIV-1 to T cells. We have also examined the role of a conserved 12 amino acid region in murine leukemia virus capsid. We demonstrate that capsid mutants which lack this conserved region, do not have a condensed core. Additionally, cell-free virion RNA isolated from these mutants have a faster mobility in an agarose gel and dissociate from a dimer to monomer ten degrees lower compared to wildtype, in a manner similar to immature RNA dimers. Thus, these mutants lack the ability to undergo virion maturation as well as RNA dimer maturation. This is the first observation suggesting a potential role of capsid in RNA dimer maturation. The studies reported in this dissertation will provide a better understanding of the nature of retroviral replication and infection.
Dang, Que, "Insights into the nature of retroviral replication and infection: Analyses of minus -strand DNA transfer, double infection, and virion and RNA dimer maturation." (2002). Graduate Theses, Dissertations, and Problem Reports. 8712.